Studies with intact cells of the eukaryotic microorganism Neurospora crassa indicate that the trans-plasma membrane electrical potential may be a primary regulator of adenylate cyclase activity in this organism. In this laboratory, methods have been developed for isolating Neurospora plasma membranes in high yield and purity, and generating a transmembrane electrical potential in inverted plasma membrane vesicles via the plasma membrane ATPase. The objectives of the proposed studies are to demonstrate in vitro that the activity of the Neurospora adenylate cyclase is indeed regulated by the membrane potential and to investigate the molecular nature of this potential new form of enzyme regulation.